Monocytes and lymphocytes as active participants in the pathogenesis of experimental shock

We investigated the role played by monocytes and lymphocytes in the pathogenesis of experimental shock. Splanchnic artery occlusion (SAO) shock was induced in anaesthetized rats by clamping splanchnic arteries for 45 min followed by reperfusion. Sham operated animals were used as controls. SAO shocked rats had a decreased survival time (80±11 min, while sham shocked rats survived more than 4 h), increased serum (248±21 U/ml) and macrophage (145±15 U/ml) levels of TNF-, enhanced myeloperoxidase (MPO) activity in the ileum (3.38±0.2 U×10^–3/g tissue), decreased number of monocytes, lymphocytes and neutrophils and a profound hypotension. In addition we found an increased expression of vascular cell adhesion molecule-1 (VCAM-1) on aortic endothelium and a reduced percentage of VLA-4 positive monocytes and lymphocytes. Inhibition of TNF- synthesis, reversed the increased endothelial expression of VCAM-1, increased the percentage of integrin VLA-4 positive leukocytes and improved monocyte, lymphocyte and neutrophil count. Furthermore a passive immunization with specific antibodies raised against VCAM-1 (2 mg/kg, i.v. 3 h before SAO) increased survival, reduced MPO activity in the ileum (0.034±0.04 U×10^–3/g tissue) and improved mean arterial blood pressure. Our data suggest that monocytes and lymphocytes participate in the pathogenesis of splanchnic ischaemia-reperfusion injury and may amplify the adhesion of neutrophils to peripheral tissues.by I. Ahnfelt Rønne     

Experimental ischaemia-reperfusion injury induces vascular endothelial growth factor expression in the rat testis

Testicular torsion causes ischaemia-reperfusion (I-R) injury of testis and might lead to male infertility. Its injury initiates a pathophysiological cascade, including an activation of inflammatory cytokines and generation of nitric oxide and other reactive oxygen species. Vascular endothelial growth factor (VEGF) mediates angiogenesis and promotes endothelial cell survival. The aim of our study was to investigate the time course expression of VEGF, VEGF-receptor (R)1, VEGF-R2, nitric oxide synthases (NOS) in experimental I-R injury of rat testis. In torsion side testis, the expression of VEGF protein and mRNA significantly increased in a time-dependent manner ( P  < 0.001 and P  < 0.001, respectively). Although the expression of VEGF-R1 mRNA was increased in a similar way ( P  < 0.001), VEGF-R2 mRNA expression was not detected. In immunohistochemistry, the increase in VEGF protein staining was observed in testicular vascular endothelial cells and germ cells at 24 h after reperfusion. Significant activation of inducible NOS and endothelial NOS was investigated at 12 and 24 h after reperfusion ( P  < 0.01 and P  < 0.001, respectively). This is the first report to show the time course expression of VEGF in experimental I-R rat testis.     

Pioglitazone,a Specific Ligand of the Peroxisome Proliferator-Activated Receptor Gamma Reduces Gastric Mucosal Injury Induced by Ischaemia/Reperfusion in Rat

Background: The peroxisome proliferator-activated receptor gamma (PPARγ) is a ligand-dependent nuclear receptor that has been implicated in the control of metabolism and numerous cellular processes, including cell cycle control, carcinogenesis, and inflammation. The present study was designed to investigate the effect of the specific PPARγ ligand, pioglitazone, on the mucosal lesions induced by ischaemia and reperfusion (I/R) in rats. Methods: I/R lesions were induced in Wistar rats by applying a small clamp to the coeliac artery for 30?min (ischaemic phase), followed by the removal of the clamp for 3?h (reperfusion phase). Vehicle (saline) or increasing doses of pioglitazone (2.5, 10, and 30?mg/kg i.g.) were given 30?min before exposure to I/R. The animals were killed immediately after the end of the reperfusion phase (time 0) and at 12 and 24?h after I/R. The area of gastric lesions was measured by planimetry, and the gastric blood flow was determined by the H[Formula: See Text] gas clearance method. The gastric mucosal gene expressions of PPARγ, interleukin-1beta (IL-1β), tumour necrosis factor alpha (TNF-α), leptin, cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) were examined by RT-PCR. In addition, protein expression of COX-2 and leptin was assessed by Western blot. Results: The pretreatment with pioglitazone reduced in a dose-dependent manner the mean lesion area induced by I/R, and this effect was accompanied by a significant increase in the gastric blood flow. The decrease in gastric ulcerations by pioglitazone was also observed 12 and 24?h after the I/R. The PPARγ mRNA was weakly expressed in the intact gastric mucosa, but significantly up-regulated after exposure to I/R at each time interval studied. The expression of IL-1β was not changed significantly after pioglitazone applied i.g. at doses 2.5 and 10?mg/kg, but it was down-regulated at the dose 30?mg/kg. TNFα mRNA was strongly increased after the exposure to I/R, but it was down-regulated after pioglitazone pretreatment. In contrast, both leptin and COX-2 mRNA and protein expression were increased in the gastric mucosa after exposure to I/R. The pretreatment with pioglitazone caused a significant up-regulation of mRNA and protein expression of leptin, reaching its peak at the dose 30?mg/kg i.g. In contrast, COX-2 expression did not change significantly after the 2.5 and 10?mg/kg of pioglitazone, but it significantly decreased after pioglitazone at dose 30?mg/kg given to rats before exposure to I/R. Conclusions: Pioglitazone reduces the acute erosions and deeper gastric lesions induced by I/R. The beneficial effect of this PPARγ ligand on I/R-induced gastric damage may be due to its anti-inflammatory properties, especially to the reduction in TNF-α expression and to up-regulation of leptin mRNA in the gastric mucosa. The inhibition of COX-2 expression by pioglitazone may reflect the anti-inflammatory properties of this compound.     

Cardioprotection by breathing hyperoxic gas—relation to oxygen concentration and exposure time in rats and mice

Objectives: Breathing a hyperoxic gas (≥95% O₂) protects against ischaemia-reperfusion injury in rat and mouse hearts. The present study investigated how oxygen concentration and duration of hyperoxic exposure influenced cardioprotection, and whether hyperoxia might induce delayed cardioprotection (after 24 h). Methods: Animals were kept in normal air or in a hyperoxic environment, and their hearts were isolated and Langendorff-perfused immediately or 24 h thereafter. Global ischaemia was induced for 25 min in rats and 40 min in mice, followed by 60 min of reperfusion. Infarct size was determined by triphenyl tetrazolium chloride staining. Results: In rats exposure to ≥95, 80, and 60%, but not to 40% of oxygen immediately before heart isolation and perfusion improved postischaemic functional recovery. Eighty or more percent of oxygen also reduced infarct size. A preconditioning-like effect could be evoked by 60 or 180 min of hyperoxia, giving both immediate and delayed protection. In the mouse heart protection could be induced by pretreatment for 15 or 30, but not by 60 min with ≥95% oxygen. The protective effect of hyperoxia in mice could be evoked in the immediate model only. Conclusions: Hyperoxia protects the isolated rat and mouse heart against ischaemia-reperfusion injury, but some species-different responses exist. The protection depends on both oxygen concentration in inspired air, and duration of hyperoxic exposure.     

药物后处理对心肌保护作用机制的研究进展

缺血后处理是在心肌遭受致命性的再灌注损伤的初始阶段,所给予的一种可控的、短暂的、反复间断的缺血/再灌注处理,这样可以明显减轻心肌的缺血/再灌注损伤。近年来,人们又日益深入地研究一种新的心肌保护策略:药物后处理。现对药物后处理对缺血/再灌注心肌保护作用机制的研究进展进行综述。     

Machine perfusion of the liver: Which is the best technique to mitigate ischaemia-reperfusion injury?

Longstanding research describes the mechanisms whereby the restoration of blood flow and reoxygenation(reperfusion) aggravates the ischaemic injury caused by a period of anoxia to a donor liver. This phenomenon, called ischaemia-reperfusion injury(IRI), leads to parenchymal cell death,microcirculatory failure, and inflammatory immune response. Clinically, IRI is the main factor responsible for the occurrence of posttransplant graft dysfunction and ischaemic-type biliary lesions. While extended criteria donor livers are more vulnerable to IRI, their utilisation is required to address the shortfall in donor organs. Thus, the mitigation of IRI should drive the setting of a new benchmark for marginal organ preservation. Herein, strategies incorporating different modalities of machine perfusion of the liver to alleviate IRI are discussed in conjunction with advantages and disadvantages of individual protocols.Techniques leading to reperfusion of the liver during machine perfusion(in situ normothermic regional perfusion and ex situ normothermic machine perfusion)may mitigate IRI by shortening the ischaemic period of the organs. This benefit potentially escalates from the minimum level, obtained following just partial alleviation of the ischaemic period, to the maximum level, which can be potentially achieved with ischaemia-free organ transplantation. Techniques that do not lead to reperfusion of the liver during machine perfusion(hypothermic,subnormothermic, and controlled-oxygenated rewarming) optimise mitochondrial oxidative function and replenish cellular energy stores, thereby lowering reactive oxygen species production as well as the activation ofdownstream inflammatory pathways during reperfusion. Further mechanistic insights into IRI may guide the development of donor-specific protocols of machine perfusion on the basis of the limitations of individual categories of extended criteria donor organs.     

Ischaemic post-conditioning protects lung from ischaemia-reperfusion injury by up-regulation of haeme oxygenase-1

Objective

The emergence of ischaemic post-conditioning (IPO) provides a potential method for experimentally and clinically attenuating various types of organ injuries. There has been little work, however, examining its effects in the setting of lung ischaemia reperfusion (IR). The stress protein, haeme oxygenase-1 (HO-1), has been found to exert a potent, protective role in a variety of lung injury models. In this study, we hypothesised that the induction of HO-1 by IPO plays a protective role against the deleterious effects of IR in the lung.

Methods

Anaesthetised and mechanically ventilated adult Sprague-Dawley rats were randomly assigned to one of the following groups (n = 8 each): the sham-operated control group, the IR group (40 min of left-lung ischaemia and 105 min of reperfusion), the IPO group (three successive cycles of 30-s reperfusion per 30-s occlusion before restoring full perfusion) and the ZnPPIX + IPO group (ZnPPIX, an inhibitor of HO-1, was injected intra-peritoneally at 20 mg kg⁻¹ 24 h prior to the experiment and the rest of the procedures were similar to that of the IPO group). Lung injury was assessed by arterial blood gas analysis, wet-to-dry lung weight ratio and tissue histological changes. The extent of lipid peroxidation was determined by measuring plasma levels of malondialdehyde (MDA) production. Expression of HO-1 was determined by immunohistochemistry.

Results

Lung IR resulted in a significant reduction of PaO₂ (data in IR, P < 0.05 vs. data in sham) and increase of lung wet-to-dry weight ratio, accompanied with increased MDA production and severe lung pathological morphological changes as well as a compensatory increase in HO-1 protein expression, as compared with sham (All P < 0.05). IPO markedly attenuated all the above pathological changes seen in the IR group and further increased HO-1 expression. Treatment with ZnPPIX abolished all the protective effects of post-conditioning.

Conclusion

It may be concluded that IPO protects IR-induced lung injury via induction of HO-1.     

Remote Ischaemic Pre-Conditioning Reduces Intestinal Ischaemia Reperfusion Injury in a Newborn Rat

ObjectiveRemote ischaemic conditioning (RIC) has been shown to reduce ischaemia-reperfusion injury(IRI) in multiple organ systems. IRI is seen in multiple bowel pathologies in the newborn, including NEC. We investigated the potential of RIC as a novel therapy for various intestinal pathologies in the newborn.MethodsWe used an established intestinal IRI model in rat pups which results in similar intestinal injury to necrotising enterocolitis (NEC). Animals were randomly allocated to IRI only(n = 14), IRI + RIC(n = 13) or sham laparotomy(n = 10). The macroscopic extent of intestinal injury is reported as a percentage of total small bowel. Injury severity was measured using Chiu-Park scoring. Neutrophil infiltration/activation was assayed by myeloperoxidase activity. Immunohistochemistry was used to assess the expression of hypoxia-inducible factor alpha (HIF-1α). Data are median (interquartile range).ResultsAnimals that underwent RIC showed a decreased extent of macroscopic injury from 100%(85–100%) in the IRI only group to 58%(15–84%, p = 0.003) in the IRI + RIC group. Microscopic injury score was significantly lower in animals that underwent RIC compared to IRI alone (3.5[1.25–5] vs 5.5[4–6], p = 0.014). Intestinal myeloperoxidase activity in animals exposed to IRI was 3.4 mU/mg of tissue (2.5–3.7) and 2.1 mU/mg(1.5–2.8) in the IRI + RIC group(p = 0.047). HIF-1α expression showed a non-significant trend towards reduced expression in the IRI + RIC group.ConclusionsRIC reduces the extent and severity of bowel injury in this animal model, supporting the hypothesis that RIC has therapeutic potential for intestinal diseases in the newborn.